The most frequent cause of acute myeloid leukemia (AML) is the 8;21 translocation [t(8;21)], which results in transcriptional dysregulation. This translocation generates an AML1-ETO fusion protein by joining part of the AML1/RUNX1 transcription factor to a nearly complete ETO protein, the prototypical member of a family of transcriptional corepressors. The long-term goal of this proposal is to learn how to selectively interfere with AML1-ETO activity, and thereby reverse the leukemogenic state. The immediate goal of this application is to understand the mechanisms by which AML1-ETO disrupts the normal transcriptional program. Aberrant expression of AML1-ETO is the pathological cause of t(8;21) AML. Phenotypic differences between the AML1 knockout and the AML1-ETO knock-in mouse models indicate that AML1-ETO has other activities besides deregulation of AML1 functions. Although it is now clear that AML1-ETO interferes with multiple cellular events involved in hematopoietic cell self-renewal, differentiation, and apoptosis, it remains unclear how AML1-ETO deregulates these pathways. Recently, Dr. Zhang discovered a molecular interaction between AML1-ETO and the class I helix-loop-helix transcriptional factors known as E proteins. Through the ETO domain, AML1-ETO aberrantly represses E protein-mediated transcription. E proteins have tumor- suppressor activities that are frequently inactivated in cancers. That is, they promote apoptosis and control hematopoietic cell differentiation. The leukemogenic potential of AML1-ETO is consistent with its inhibition of E protein functions related to both tumor suppression and regulation of cell differentiation. Dr. Zhang's preliminary studies show (i) that the ETO domains involved in repressing E protein- dependent transcription correlate with those involved in the leukemogenic activities of AML1-ETO; and (ii) that repression of E protein-dependent transcription by AML1-ETO involves not only chromatin-dependent inhibition, but also direct inhibition of the RNA polymerase II transcription complex. These findings led to the central hypothesis that AML1-ETO must repress both the chromatin-dependent and chromatin- independent transcription mediated by E proteins to allow for leukemogenesis. The hypothesis will be tested through the following two aims: (Aim 1) To define the mechanisms by which AML1-ETO represses E protein-dependent transcription at the level of chromatin as well as at the level of basal transcription machinery; and (Aim 2) To determine the extent to which inactivation of E proteins contributes to AML1-ETO leukemogenic function, and to elucidate the molecular pathways associated with E proteins in t(8;21) leukemic cells. A better understanding of the molecular mechanisms underlying t(8;21) AML, and the aberrant functions of proteins involved in leukemogenesis should lead to the identification of new therapeutic targets and strategies for treatment of AML.